RESUMO
Plants are foundational for global ecological and economic systems, but most plant proteins remain uncharacterized. Protein interaction networks often suggest protein functions and open new avenues to characterize genes and proteins. We therefore systematically determined protein complexes from 13 plant species of scientific and agricultural importance, greatly expanding the known repertoire of stable protein complexes in plants. By using co-fractionation mass spectrometry, we recovered known complexes, confirmed complexes predicted to occur in plants, and identified previously unknown interactions conserved over 1.1 billion years of green plant evolution. Several novel complexes are involved in vernalization and pathogen defense, traits critical for agriculture. We also observed plant analogs of animal complexes with distinct molecular assemblies, including a megadalton-scale tRNA multi-synthetase complex. The resulting map offers a cross-species view of conserved, stable protein assemblies shared across plant cells and provides a mechanistic, biochemical framework for interpreting plant genetics and mutant phenotypes.
Assuntos
Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Mapas de Interação de Proteínas/fisiologia , Espectrometria de Massas/métodos , Plantas/genética , Plantas/metabolismo , Mapeamento de Interação de Proteínas/métodos , Proteômica/métodosRESUMO
The discovery and application of genome editing introduced a new era of plant breeding by giving researchers efficient tools for the precise engineering of crop genomes1. Here we demonstrate the power of genome editing for engineering broad-spectrum disease resistance in rice (Oryza sativa). We first isolated a lesion mimic mutant (LMM) from a mutagenized rice population. We then demonstrated that a 29-base-pair deletion in a gene we named RESISTANCE TO BLAST1 (RBL1) caused broad-spectrum disease resistance and showed that this mutation caused an approximately 20-fold reduction in yield. RBL1 encodes a cytidine diphosphate diacylglycerol synthase that is required for phospholipid biosynthesis2. Mutation of RBL1 results in reduced levels of phosphatidylinositol and its derivative phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). In rice, PtdIns(4,5)P2 is enriched in cellular structures that are specifically associated with effector secretion and fungal infection, suggesting that it has a role as a disease-susceptibility factor3. By using targeted genome editing, we obtained an allele of RBL1, named RBL1Δ12, which confers broad-spectrum disease resistance but does not decrease yield in a model rice variety, as assessed in small-scale field trials. Our study has demonstrated the benefits of editing an LMM gene, a strategy relevant to diverse LMM genes and crops.
Assuntos
Diacilglicerol Colinofosfotransferase , Resistência à Doença , Edição de Genes , Oryza , Melhoramento Vegetal , Doenças das Plantas , Resistência à Doença/genética , Edição de Genes/métodos , Genoma de Planta/genética , Oryza/enzimologia , Oryza/genética , Oryza/microbiologia , Fosfatidilinositóis/metabolismo , Melhoramento Vegetal/métodos , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Alelos , Fosfatidilinositol 4,5-Difosfato/metabolismo , Diacilglicerol Colinofosfotransferase/genética , Diacilglicerol Colinofosfotransferase/metabolismoRESUMO
Climate change is a defining challenge of the 21st century, and this decade is a critical time for action to mitigate the worst effects on human populations and ecosystems. Plant science can play an important role in developing crops with enhanced resilience to harsh conditions (e.g. heat, drought, salt stress, flooding, disease outbreaks) and engineering efficient carbon-capturing and carbon-sequestering plants. Here, we present examples of research being conducted in these areas and discuss challenges and open questions as a call to action for the plant science community.
Assuntos
Mudança Climática , Ecossistema , Humanos , Produtos Agrícolas , Carbono , SecasRESUMO
Climate change is affecting the types of plant varieties we can cultivate, as well as how and where we can do so. A new collection of articles explores the twin challenges of engineering plants for resilience to climate change and enhancing their carbon-capture potential.
Assuntos
Mudança Climática , Plantas , Plantas/genética , CarbonoRESUMO
Root-knot nematodes (Meloidogyne spp.) are highly evolved obligate parasites threatening global food security. These parasites have a remarkable ability to establish elaborate feeding sites in roots, which are their only source of nutrients throughout their life cycle. A wide range of nematode effectors have been implicated in modulation of host pathways for defense suppression and/or feeding site development. Plants produce a diverse array of peptide hormones including PLANT PEPTIDE CONTAINING SULFATED TYROSINE (PSY)-family peptides, which promote root growth via cell expansion and proliferation. A sulfated PSY-like peptide RaxX (required for activation of XA21 mediated immunity X) produced by the biotrophic bacterial pathogen (Xanthomonas oryzae pv. oryzae) has been previously shown to contribute to bacterial virulence. Here, we report the identification of genes from root-knot nematodes predicted to encode PSY-like peptides (MigPSYs) with high sequence similarity to both bacterial RaxX and plant PSYs. Synthetic sulfated peptides corresponding to predicted MigPSYs stimulate root growth in Arabidopsis. MigPSY transcript levels are highest early in the infection cycle. Downregulation of MigPSY gene expression reduces root galling and egg production, suggesting that the MigPSYs serve as nematode virulence factors. Together, these results indicate that nematodes and bacteria exploit similar sulfated peptides to hijack plant developmental signaling pathways to facilitate parasitism.
Assuntos
Arabidopsis , Nematoides , Parasitos , Tylenchoidea , Animais , Plantas , Peptídeos , Transdução de Sinais , Tirosina , Doenças das Plantas/microbiologia , Tylenchoidea/genética , Raízes de PlantasRESUMO
Conventional methods of DNA sequence insertion into plants, using Agrobacterium-mediated transformation or microprojectile bombardment, result in the integration of the DNA at random sites in the genome. These plants may exhibit altered agronomic traits as a consequence of disruption or silencing of genes that serve a critical function. Also, genes of interest inserted at random sites are often not expressed at the desired level. For these reasons, targeted DNA insertion at suitable genomic sites in plants is a desirable alternative. In this paper we review approaches of targeted DNA insertion in plant genomes, discuss current technical challenges, and describe promising applications of targeted DNA insertion for crop genetic improvement.
Assuntos
Produtos Agrícolas/genética , DNA de Plantas/genética , Técnicas de Transferência de Genes , Genoma de Planta , Plantas Geneticamente Modificadas/genética , Transformação Genética , AgrobacteriumRESUMO
Parent-of-origin-dependent gene expression in mammals and flowering plants results from differing chromatin imprints (genomic imprinting) between maternally and paternally inherited alleles. Imprinted gene expression in the endosperm of seeds is associated with localized hypomethylation of maternally but not paternally inherited DNA, with certain small RNAs also displaying parent-of-origin-specific expression. To understand the evolution of imprinting mechanisms in Oryza sativa (rice), we analyzed imprinting divergence among four cultivars that span both japonica and indica subspecies: Nipponbare, Kitaake, 93-11, and IR64. Most imprinted genes are imprinted across cultivars and enriched for functions in chromatin and transcriptional regulation, development, and signaling. However, 4 to 11% of imprinted genes display divergent imprinting. Analyses of DNA methylation and small RNAs revealed that endosperm-specific 24-nt small RNA-producing loci show weak RNA-directed DNA methylation, frequently overlap genes, and are imprinted four times more often than genes. However, imprinting divergence most often correlated with local DNA methylation epimutations (9 of 17 assessable loci), which were largely stable within subspecies. Small insertion/deletion events and transposable element insertions accompanied 4 of the 9 locally epimutated loci and associated with imprinting divergence at another 4 of the remaining 8 loci. Correlating epigenetic and genetic variation occurred at key regulatory regions-the promoter and transcription start site of maternally biased genes, and the promoter and gene body of paternally biased genes. Our results reinforce models for the role of maternal-specific DNA hypomethylation in imprinting of both maternally and paternally biased genes, and highlight the role of transposition and epimutation in rice imprinting evolution.
Assuntos
Endosperma/genética , Evolução Molecular , Impressão Genômica , Oryza/genética , Metilação de DNA , Elementos de DNA Transponíveis , Epigenômica , Regulação da Expressão Gênica de Plantas , Mutação , Oryza/classificação , Oryza/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismoRESUMO
Early root growth is critical for plant establishment and survival. We have identified a molecular pathway required for helical root tip movement known as circumnutation. Here, we report a multiscale investigation of the regulation and function of this phenomenon. We identify key cell signaling events comprising interaction of the ethylene, cytokinin, and auxin hormone signaling pathways. We identify the gene Oryza sativa histidine kinase-1 (HK1) as well as the auxin influx carrier gene OsAUX1 as essential regulators of this process in rice. Robophysical modeling and growth challenge experiments indicate circumnutation is critical for seedling establishment in rocky soil, consistent with the long-standing hypothesis that root circumnutation facilitates growth past obstacles. Thus, the integration of robotics, physics, and biology has elucidated the functional importance of root circumnutation and uncovered the molecular mechanisms underlying its regulation.
Assuntos
Regulação da Expressão Gênica de Plantas , Histidina Quinase/metabolismo , Ácidos Indolacéticos/farmacologia , Oryza/crescimento & desenvolvimento , Proteínas de Plantas/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Solo/química , Transporte Biológico , Citocininas/metabolismo , Histidina Quinase/genética , Oryza/efeitos dos fármacos , Oryza/genética , Oryza/metabolismo , Reguladores de Crescimento de Plantas/farmacologia , Proteínas de Plantas/genética , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/metabolismoRESUMO
Tyrosine sulfation, a post-translational modification, can determine and often enhance protein-protein interaction specificity. Sulfotyrosyl residues (sTyrs) are formed by the enzyme tyrosyl-protein sulfotransferase during protein maturation in the Golgi apparatus and most often occur singly or as a cluster within a six-residue span. With both negative charge and aromatic character, sTyr facilitates numerous atomic contacts as visualized in binding interface structural models, thus there is no discernible binding site consensus. Found exclusively in secreted proteins, in this review, we discuss the four broad sequence contexts in which sTyr has been observed: first, a solitary sTyr has been shown to be critical for diverse high-affinity interactions, such as between peptide hormones and their receptors, in both plants and animals. Second, sTyr clusters within structurally flexible anionic segments are essential for a variety of cellular processes, including coreceptor binding to the HIV-1 envelope spike protein during virus entry, chemokine interactions with receptors, and leukocyte rolling cell adhesion. Third, a subcategory of sTyr clusters is found in conserved acidic sequences termed hirudin-like motifs that enable proteins to interact with thrombin; consequently, many proven and potential therapeutic proteins derived from blood-consuming invertebrates depend on sTyrs for their activity. Finally, several proteins that interact with collagen or similar proteins contain one or more sTyrs within an acidic residue array. Refined methods to direct sTyr incorporation in peptides synthesized both in vitro and in vivo, together with continued advances in mass spectrometry and affinity detection, promise to accelerate discoveries of sTyr occurrence and function.
Assuntos
Peptídeos , Domínios e Motivos de Interação entre Proteínas , Tirosina , Animais , Peptídeos/química , Processamento de Proteína Pós-Traducional , Proteínas/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
The rice immune receptor XA21 confers resistance to Xanthomonas oryzae pv. oryzae (Xoo), and upon recognition of the RaxX21-sY peptide produced by Xoo, XA21 activates the plant immune response. Here we screened 21 000 mutant plants expressing XA21 to identify components involved in this response, and reported here the identification of a rice mutant, sxi4, which is susceptible to Xoo. The sxi4 mutant carries a 32-kb translocation from chromosome 3 onto chromosome 7 and displays an elevated level of DCL2a transcript, encoding a Dicer-like protein. Silencing of DCL2a in the sxi4 genetic background restores resistance to Xoo. RaxX21-sY peptide-treated leaves of sxi4 retain the hallmarks of XA21-mediated immune response. However, WRKY45-1, a known negative regulator of rice resistance to Xoo, is induced in the sxi4 mutant in response to RaxX21-sY peptide treatment. A CRISPR knockout of a short interfering RNA (TE-siRNA815) in the intron of WRKY45-1 restores the resistance phenotype in sxi4. These results suggest a model where DCL2a accumulation negatively regulates XA21-mediated immunity by altering the processing of TE-siRNA815.
Assuntos
Oryza , Xanthomonas , Oryza/metabolismo , Peptídeos/metabolismo , Fenótipo , Doenças das Plantas/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Serina-Treonina Quinases/genética , Xanthomonas/metabolismoRESUMO
Systematic mappings of protein interactome networks have provided invaluable functional information for numerous model organisms. Here we develop PCR-mediated Linkage of barcoded Adapters To nucleic acid Elements for sequencing (PLATE-seq) that serves as a general tool to rapidly sequence thousands of DNA elements. We validate its utility by generating the ORFeome for Oryza sativa covering 2,300 genes and constructing a high-quality protein-protein interactome map consisting of 322 interactions between 289 proteins, expanding the known interactions in rice by roughly 50%. Our work paves the way for high-throughput profiling of protein-protein interactions in a wide range of organisms.
Assuntos
Fases de Leitura Aberta/genética , Oryza/genética , Mapeamento de Interação de Proteínas/métodos , Mapas de Interação de Proteínas/genética , Análise de Sequência de DNA/métodos , Biologia Computacional/métodos , DNA de Plantas/genética , Bases de Dados Genéticas , Genoma de Planta/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodosRESUMO
The rice immune receptor XA21 is activated by the sulfated microbial peptide required for activation of XA21-mediated immunity X (RaxX) produced by Xanthomonas oryzae pv. oryzae (Xoo). Mutational studies and targeted proteomics revealed that the RaxX precursor peptide (proRaxX) is processed and secreted by the protease/transporter RaxB, the function of which can be partially fulfilled by a noncognate peptidase-containing transporter component B (PctB). proRaxX is cleaved at a Gly-Gly motif, yielding a mature peptide that retains the necessary elements for RaxX function as an immunogen and host peptide hormone mimic. These results indicate that RaxX is a prokaryotic member of a previously unclassified and understudied group of eukaryotic tyrosine sulfated ribosomally synthesized, posttranslationally modified peptides (RiPPs). We further demonstrate that sulfated RaxX directly binds XA21 with high affinity. This work reveals a complete, previously uncharacterized biological process: bacterial RiPP biosynthesis, secretion, binding to a eukaryotic receptor, and triggering of a robust host immune response.
Assuntos
Proteínas de Bactérias/metabolismo , Peptídeo Hidrolases/metabolismo , Peptídeos/metabolismo , Proteínas de Plantas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transportadores de Cassetes de Ligação de ATP/química , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Genes Bacterianos/genética , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Redes e Vias Metabólicas/genética , Oryza/imunologia , Oryza/metabolismo , Oryza/microbiologia , Peptídeo Hidrolases/química , Peptídeo Hidrolases/genética , Peptídeos/química , Peptídeos/genética , Proteínas de Plantas/química , Proteínas de Plantas/imunologia , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/imunologia , Xanthomonas/genética , Xanthomonas/metabolismo , Xanthomonas/patogenicidadeRESUMO
Upon encountering a susceptible plant host, a bacterial pathogen expresses specific virulence factors. For example, in planta, the Xanthomonas HrpX protein activates transcription of roughly 150 genes encoding components of the type III secretion system or its translocated effectors, as well as other secreted proteins implicated in pathogenesis. Here, we show that X. oryzae pv. oryzae growth in planta or in HrpX-inducing XOM2 media resulted in HrpX-dependent transcription of the raxX and raxST genes that control production of the RaxX sulfopeptide, exported through a type I secretion system. The RaxX protein is required for activation of XA21-mediated immunity in Xa21+ rice lines. We identified potential plant-inducible promoter elements upstream of the likely 5' ends of the raxX and raxST transcripts. Deletions and nucleotide substitutions confirmed that these elements are required for HrpX-dependent expression of raxX and raxST. We conclude that raxX-raxST gene expression is induced by HrpX during growth in planta and, therefore, is coordinately expressed with other genes required for pathogenesis.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
Assuntos
Oryza , Doenças das Plantas , Imunidade Vegetal , Xanthomonas , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Oryza/metabolismo , Oryza/microbiologia , Doenças das Plantas/imunologia , Doenças das Plantas/microbiologia , Sistemas de Secreção Tipo III/genética , Sistemas de Secreção Tipo III/metabolismo , Xanthomonas/patogenicidadeRESUMO
BACKGROUND: Lignin deposited in plant cell walls negatively affects biomass conversion into advanced bioproducts. There is therefore a strong interest in developing bioenergy crops with reduced lignin content or altered lignin structures. Another desired trait for bioenergy crops is the ability to accumulate novel bioproducts, which would enhance the development of economically sustainable biorefineries. As previously demonstrated in the model plant Arabidopsis, expression of a 3-dehydroshikimate dehydratase in plants offers the potential for decreasing lignin content and overproducing a value-added metabolic coproduct (i.e., protocatechuate) suitable for biological upgrading. RESULTS: The 3-dehydroshikimate dehydratase QsuB from Corynebacterium glutamicum was expressed in the bioenergy crop switchgrass (Panicum virgatum L.) using the stem-specific promoter of an O-methyltransferase gene (pShOMT) from sugarcane. The activity of pShOMT was validated in switchgrass after observation in-situ of beta-glucuronidase (GUS) activity in stem nodes of plants carrying a pShOMT::GUS fusion construct. Under controlled growth conditions, engineered switchgrass lines containing a pShOMT::QsuB construct showed reductions of lignin content, improvements of biomass saccharification efficiency, and accumulated higher amount of protocatechuate compared to control plants. Attempts to generate transgenic switchgrass lines carrying the QsuB gene under the control of the constitutive promoter pZmUbi-1 were unsuccessful, suggesting possible toxicity issues associated with ectopic QsuB expression during the plant regeneration process. CONCLUSION: This study validates the transfer of the QsuB engineering approach from a model plant to switchgrass. We have demonstrated altered expression of two important traits: lignin content and accumulation of a co-product. We found that the choice of promoter to drive QsuB expression should be carefully considered when deploying this strategy to other bioenergy crops. Field-testing of engineered QsuB switchgrass are in progress to assess the performance of the introduced traits and agronomic performances of the transgenic plants.
Assuntos
Corynebacterium/enzimologia , Hidroliases/metabolismo , Lignina/biossíntese , Panicum/genética , Regiões Promotoras Genéticas/genética , Saccharum/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biomassa , Parede Celular/metabolismo , Corynebacterium/genética , Regulação da Expressão Gênica de Plantas , Genes Reporter , Hidroliases/genética , Lignina/análise , Metiltransferases/genética , Especificidade de Órgãos , Panicum/crescimento & desenvolvimento , Panicum/metabolismo , Proteínas de Plantas/genética , Caules de Planta/enzimologia , Caules de Planta/genética , Plantas Geneticamente Modificadas , Saccharum/enzimologiaRESUMO
Crops carrying broad-spectrum resistance loci provide an effective strategy for controlling infectious disease because these loci typically confer resistance to diverse races of a pathogen or even multiple species of pathogens. Despite their importance, only a few crop broad-spectrum resistance loci have been reported. Here, we report the identification and characterization of the rice bsr-k1 (broad-spectrum resistance Kitaake-1) mutant, which confers broad-spectrum resistance against Magnaporthe oryzae and Xanthomonas oryzae pv oryzae with no major penalty on key agronomic traits. Map-based cloning reveals that Bsr-k1 encodes a tetratricopeptide repeats (TPRs)-containing protein, which binds to mRNAs of multiple OsPAL (OsPAL1-7) genes and promotes their turnover. Loss of function of the Bsr-k1 gene leads to accumulation of OsPAL1-7 mRNAs in the bsr-k1 mutant. Furthermore, overexpression of OsPAL1 in wild-type rice TP309 confers resistance to M. oryzae, supporting the role of OsPAL1 Our discovery of the bsr-k1 allele constitutes a significant conceptual advancement and provides a valuable tool for breeding broad-spectrum resistant rice.
Assuntos
Oryza/fisiologia , Doenças das Plantas/genética , Proteínas de Plantas/genética , Proteínas de Ligação a RNA/genética , Citoplasma/metabolismo , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Magnaporthe/patogenicidade , Mutação , Oryza/genética , Oryza/microbiologia , Doenças das Plantas/microbiologia , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Domínios Proteicos , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Sequências Repetitivas de Aminoácidos , Xanthomonas/patogenicidadeRESUMO
The availability of a whole-genome sequenced mutant population and the cataloging of mutations of each line at a single-nucleotide resolution facilitate functional genomic analysis. To this end, we generated and sequenced a fast-neutron-induced mutant population in the model rice cultivar Kitaake (Oryza sativa ssp japonica), which completes its life cycle in 9 weeks. We sequenced 1504 mutant lines at 45-fold coverage and identified 91,513 mutations affecting 32,307 genes, i.e., 58% of all rice genes. We detected an average of 61 mutations per line. Mutation types include single-base substitutions, deletions, insertions, inversions, translocations, and tandem duplications. We observed a high proportion of loss-of-function mutations. We identified an inversion affecting a single gene as the causative mutation for the short-grain phenotype in one mutant line. This result reveals the usefulness of the resource for efficient, cost-effective identification of genes conferring specific phenotypes. To facilitate public access to this genetic resource, we established an open access database called KitBase that provides access to sequence data and seed stocks. This population complements other available mutant collections and gene-editing technologies. This work demonstrates how inexpensive next-generation sequencing can be applied to generate a high-density catalog of mutations.
Assuntos
Genoma de Planta/genética , Genômica/métodos , Oryza/genética , DNA de Plantas/genética , Mutação/genética , Análise de Sequência de DNARESUMO
BACKGROUND: The availability of thousands of complete rice genome sequences from diverse varieties and accessions has laid the foundation for in-depth exploration of the rice genome. One drawback to these collections is that most of these rice varieties have long life cycles, and/or low transformation efficiencies, which limits their usefulness as model organisms for functional genomics studies. In contrast, the rice variety Kitaake has a rapid life cycle (9 weeks seed to seed) and is easy to transform and propagate. For these reasons, Kitaake has emerged as a model for studies of diverse monocotyledonous species. RESULTS: Here, we report the de novo genome sequencing and analysis of Oryza sativa ssp. japonica variety KitaakeX, a Kitaake plant carrying the rice XA21 immune receptor. Our KitaakeX sequence assembly contains 377.6 Mb, consisting of 33 scaffolds (476 contigs) with a contig N50 of 1.4 Mb. Complementing the assembly are detailed gene annotations of 35,594 protein coding genes. We identified 331,335 genomic variations between KitaakeX and Nipponbare (ssp. japonica), and 2,785,991 variations between KitaakeX and Zhenshan97 (ssp. indica). We also compared Kitaake resequencing reads to the KitaakeX assembly and identified 219 small variations. The high-quality genome of the model rice plant KitaakeX will accelerate rice functional genomics. CONCLUSIONS: The high quality, de novo assembly of the KitaakeX genome will serve as a useful reference genome for rice and will accelerate functional genomics studies of rice and other species.
Assuntos
Genoma de Planta , Genômica , Oryza/genética , Sequenciamento Completo do Genoma , Biologia Computacional/métodos , Variação Genética , Genômica/métodos , Anotação de Sequência Molecular , Oryza/classificação , FenótipoRESUMO
[This corrects the article DOI: 10.1371/journal.pgen.1006049.].